Antimicrobial activity of 3-O-Acyl-(-)-epicatechin and 3-O-Acyl-(+)- catechin derivatives

Article abstract of DOI:10.1055/s-2004-818923

As an exploratory investigation of antimicrobial promoting compounds, 3-O-acyl-(-)-epicatechins and 3-O-acyl-(+)-catechins possessing various aromatic groups and aliphatic chains of varying length from C4 to C16 for increasing lipophilicity were synthesiz

Full text of DOI:10.1055/s-2004-818923

Antimicrobial Activity of
-O-Acyl-ꢀ±)-epicatechin and 3-O-
Acyl-ꢀ+)-catechin derivatives
3
1
1
1
1
Ki Duk Park , Yoon Sun Park , Sung Jin Cho , Won Suck Sun ,
2
2
1
Sung Han Kim , Do Hyun Jung , Jung Han Kim
Abstract
As an exploratory investigation of antimicrobial promoting com-
pounds, 3-O-acyl-'±)-epicatechins and 3-O-acyl-'+)-catechins
possessing various aromatic groups and aliphatic chains of vary-
ing length from C4 to C16 for increasing lipophilicity were syn-
thesized and tested for antimicrobial activities against Gram-po-
sitive, Gram-negative bacteria and fungi. The '±)-epicatechin and
'+)-catechin derivatives comprised of aromatic groups increased
activity and derivatives with acyl chain groups of carbon atoms
in the close vicinity of C8 to C10 showed strong antimicrobial ac-
tivity 'MIC = 2±8 mg/ml) against Gram-positive bacteria and
weak activity against fungi. However, the activity decreased
when the carbon chain length of the substituents was too short
'C4 to C6) or too long 'C16). These results suggest that the pres-
ence of lipophilic substituents with moderate sizes might be cru-
cial for the optimal antimicrobial activity.
Tea, Camellia sinensis, has recently attracted much attention with
respect to the beneficial biological activities of its compounds ca-
techins, including antimutagenic, antibacterial, hypocholestero-
lenic, antioxidant, antitumor and cancer preventive properties
[
1], [2], [3], [4], [5], [6], [7], [8], [9]. Green tea contains many
2
72
polyphenols such as the catechins, which include '±)-epigalloca-
techin 3-gallate ['±)-EGCG], '±)-epigallocatechin ['±)-EGC], '±)-
epicatechin 3-gallate ['±)-ECG] '1), '±)-epicatechin ['±)-EC] '2)
and '+)-catechin ['+)-C] '3) [10] 'Fig.1).
Using an antimicrobial assay, '±)-ECG and '±)-EGCG exhibited
better activity than '±)-EC and '±)-EGC [11]. Since '±)-ECG and
'±)-EGCG only differ structurally from '±)-EC and '±)-EGC by
the presence of a gallic acid ester on the 3-hydroxy, the gallate
group is perceived to be important for the enhanced ability of '±)-
ECG and '±)-EGCG to have antimicrobial activity.
To determine whether structural changes to the lipophilic sub-
stituents in the gallate group could enhance activity, a series of
'±)-EC derivatives and their epimer, '+)-C derivatives, were syn-
thesized as presented in Fig. 2.
Affiliation: 1 Department of Biotechnology, College of Engineering, Yonsein
University, Seoul, Korea ´ 2 Nutrex Technology, Seongnam-si, Kyounggi-do,
Korea
Correspondence: Dr. J. H. Kim ´ Department of Biotechnology ´ College of Engi-
neering ´ Yonsei University ´ 134 Shinchon-dong ´ Seoul 120±749 ´ Korea ´ Fax:
+
822-362-7265´ E-mail: junghan@yonsei.ac.kr
Received: July 7, 2003 ´ Accepted: November 9, 2003
Bibliography: Planta Med 2004; 70: 272±276 ´ ꢀ Georg Thieme Verlag Stuttgart ´
New York ´ ISSN 0032-0943 ´ DOI 10.1055/s-2004-818923

terial activities of synthesized compounds 4 ± 29 were compared
with those of '±)-ECG '1), '±)-EC '2), '+)-C '3) and the positive
control, kanamycin sulfate. Modification of the hydroxy groups
of the gallate ester by replacement of gallic acid with various aro-
matic groups without phenolic groups improved the antibacter-
ial activity against Gram-positive bacteria. The most significant
structural change leading to enhanced activity was the introduc-
tion of an aliphatic acid ester in place of the gallic acid ester of '±)-
ECG. Among the acylepicatechin derivatives, nonanoyl '8), de-
canoyl '9) and lauloyl '10) derivatives showed strong activity
against Gram-positive bacteria. For Gram-negative bacteria, the
synthesized compounds had little antibacterial activity. Addi-
tionally, the antifungal activity of derivatives was compared
with ECG '1), EC '2), C '3) as well as the positive control, ampho-
tericin B. Among the acylepicatechin derivatives, the octanoyl
'7), nonanoyl '8) and decanoyl '9) derivatives displayed moder-
ate activity against fungi. Acylcatechin derivatives showed sim-
ilar results as acylepicatechin derivatives.
In conclusion, it had been observed that structural changes to the
lipophilic substituents with moderate size on the 3-hydroxy sub-
stituent increased the antimicrobial activity, and 3-O-acyl-'±)-
epicatechin and 3-O-acyl-'±)-catechin derivatives with C ± C
Fig. 1 Chemical structures of major catechins of tea.
8
12
acyl chains '7 ± 10,20 ± 23) exhibited potent antimicrobial activ-
The structures of new acyl derivatives '4 ± 29) were determined by ity and therefore could be considered as promising candidates
1
13
NMR and MS experiments. Full signal assignment of H- and C- for novel antimicrobial agents.
NMR was carried out with various NMR techniques including
DEPT, COSY, H,C-COSY, and long-range H,C-COSY. The complete as-
1
signment of H-NMR chemical shifts and mass spectra for epicate- Material and Methods
chin derivatives with strong antimicrobial activity and catechin
derivatives with various aromatic groups are described in Table 1.
'±)-Epicatechin gallate '1), '±)-epicatechin '2) and '+)-catechin
3) were purchased from Sigma-Aldrich Chemical Co 'minimum
'
1
13
The biological activities of the compounds against a panel of mi- 98%). H- and C-NMR spectra were recorded on a Varian Mer-
273
croorganisms are summarized in Table 2a, 2b and 3. The antibac- cury '300 MHz) instrument using TMS as the internal standard
Fig. 2 Synthesis of 3-O-acyl-"±)-epicatechins "4 ± 16) and 3-O-acyl-"+)-catechins "17 ± 29).
Letter¼ Planta Med 2004; 70: 272±276

1
a
Table 1 H-NMR data and mass spectra of 3-O-acyl substituted derivatives of epicatechin "8 ± 10) and derivatives of catechin "26 ± 28)
CH ꢀCH ) CH -CO- or Ar-CO- at C -O
ESI MSb
formula
3
2
n
2
3
CH3-
-CH2-
-CH -CO- C4-Haxial
C4-Hequtorial C2-H
C3-H
Ar-H
A-ring)
Ar-H
ꢀB-ring)
OH
ꢀA, B-ring)
m/z
2
ꢀ
7
8
9
0.83
1.15 ~1.37 2.12
"m,10H) "t,2H)
1.15 ~1.38 2.13
"m,12H) "t,2H)
1.17 ~1.39 2.15
"m,14H) "t,2H)
1.15 ~1.40 2.13
"m,18H) "t,2H)
CH -O-Ar
2.58
"dd,1H)
2.88
"dd,1H)
4.95
"d,1H)
5.21
"m,1H)
5.75 ± 5.94 6.62 ± 6.86 8.72 ± 9.24
"2d,2H) "m,3H) "m,4H)
5.75 ± 5.95 6.62 ± 6.85 8.72 ± 9.25 C24H30O7 430
"2d,2H) "m,3H) "m,4H)
5.70 ± 5.92 6.64 ± 6.85 8.69 ± 9.23 C25H32O7 444
"2d,2H) "m,3H) "m,4H)
5.74 ± 5.95 6.65 ± 6.89 8.70 ± 9.24 C27H36O7 472
"2d,2H) "m,3H) "m,4H)
C
23
H
28
O
7
416
"
t,3H)
0.83
t,3H)
0.84
t,3H)
0.84
t,3H)
Ar-H
7.42 ± 7.85
m,5H)
7.03
s,2H)
7.42 ± 7.56
m,3H)
7.43 ± 7.52
m,2H)
2.59
"dd,1H)
2.89
"dd,1H)
4.94
"d,1H)
5.21
"m,1H)
"
2.61
"dd,1H)
2.92
"dd,1H)
4.93
"d,1H)
5.22
"m,1H)
"
10
2.61
"dd,1H)
2.90
"dd,1H)
4.94
"d,1H)
5.18
"m,1H)
"
3
26
27
28
29
2.59
"dd,1H)
2.89
"dd,1H)
4.94
"d,1H)
5.21
"m,1H)
5.75 ± 5.95 6.62 ± 6.85 8.72 ± 9.25 C23H18O7 394
"2d,2H) "m,3H) "m,4H)
5.70 ± 5.92 6.64 ± 6.85 8.69 ± 9.23 C25H24O10 484
"2d,2H) "m,3H) "m,4H)
5.74 ± 5.95 6.65 ± 6.89 8.70 ± 9.24
"2d,2H) "m,3H) "m,4H)
5.74 ± 5.95 6.66 ± 6.89 8.71 ± 9.22 C22H15O7F3 438
"2d,2H) "m,3H) "m,4H)
"
3.77
"m, 9H)
2.61
"dd,1H)
2.92
"dd,1H)
4.93
"d,1H)
5.22
"m,1H)
"
2.61
"dd,1H)
2.90
"dd,1H)
4.94
"d,1H)
5.18
"m,1H)
22 16 7 2
C H O F 430
"
2.60
"dd,1H)
2.91
"dd,1H)
4.94
"d,1H)
5.18
"m,1H)
"
a 1
b
H-NMR spectra were run in DMSO-d6 on a Varian Mercury-300M Hz "d from TMS).
Mass spectra were determined on a Fisons-VG platform in the positive ESI "electron spray ionization) mode.
Table 2a Antibacterial activities of 3-O-acylepicatechin derivatives "5 ± 16), and 3-O-acylcatechin derivatives "17 ± 29). Compounds 1 ± 4 "MIC
128) not shown.
>
Test organisms
MIC ꢀmg/mL)
5
6
7
8
9
10
11
12
13
14
15
16
17
2
74
Enterococcus faecalis
ATCC 29212
> 128
> 128
64
64
32
16
8
16
8
32
16
16
32
64
32
64
32
128
128
32
> 128
> 128
128
32
32
> 128
> 128
64
Staphylococcus aureus
ATCC 25923
64
32
64
32
16
32
16
4
8
2
8
32
16
32
32
32
64
Micrococcus luteus
ATCC 10240
4
Staphylococcus
epidermidis
> 128
16
8
64
128
> 128
ATCC 0155
Bacillus subtilis
ATCC 6633
> 128
> 128
> 128
> 128
> 128
128
128
32
16
8
64
64
128
64
128
128
> 128
> 128
> 128
> 128
> 128
Escherichia coli
ATCC 25922
> 128
> 128
128
> 128
> 128
128
> 128
> 128
128
> 128
> 128
> 128
64
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
128
> 128
> 128
> 128
128
> 128 > 128
> 128 > 128
Escherichia coli
ATCC 10536
Proteus mirabilis
ATCC 27853
32
32
Klebsiella pneumoniae
> 128
128
64
128
128
128
ATCC 10031
Determined after 24 hours of incubation at 37. for the bacteria. All experiments were run in triplicate. Km = kanamycin sulfate.
and DMSO-d as the solvent. The mass spectra were taken on a acid in tetrahydrofuran at room temperature for 24 h, respective-
6
Fisons-VG platform in the positive ESI mode.
ly [2]. Evaporation and chromatography were performed with
CHCl :MeOH '9:1) on a silica gel column and then with
3
3
-O-Acylepicatechins and 3-O-acylcatechins ꢀ 4 ± 29 ): '±)-Epica- CHCl :MeOH '1:2) on Sephadex LH-20 to remove residual acid
3
techin and '+)-catechin were reacted with 1.1 equiv, of straight- to give the final products 4 ± 29. The purities of compounds 4 ±
chain acid chlorides of C to C carbon atoms, as well as benzoyl, 29 were not les than 99% by ESI-MS.
4
16
3,4,5-trimethoxybenzoyl, 3,5-difluorobenzoyl and 2,4,5-trifluoro-
benzoyl chlorides in the presence of 2 equivs. of trifluoroacetic
Letter¼ Planta Med 2004; 70: 272±276

Table 2b Antibacterial activities of 3-O-acylepicatechin derivatives "5 ± 16), and 3-O-acylcatechin derivatives "17 ± 29). Compounds 1 ± 4 "MIC
128) not shown.
>
Test organisms
MIC ꢀmg/mL)
19
1
8
20
21
22
23
24
25
26
27
28
29
km
Enterococcus faecalis
ATCC 29212
> 128
128
32
64
32
32
16
32
16
16
4
16
8
8
8
2
4
32
16
16
16
64
32
16
64
128
128
64
> 128
> 128
128
64
64
32
64
32
32
32
64
32
16
8
Staphylococcus aureus
ATCC 25923
64
32
64
Micrococcus luteus
ATCC 10240
2
Staphylococcus
epidermidis
64
8
4
64
> 128
4
ATCC 0155
Bacillus subtilis
ATCC 6633
128
128
64
32
32
32
32
32
128
> 128
> 128
> 128
> 128
> 128
128
128
2
8
Escherichia coli
ATCC 25922
> 128
> 128
128
> 128
> 128
128
> 128
> 128
64
> 128
> 128
32
> 128
> 128
32
> 128
> 128
64
> 128
> 128
> 128
> 64
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
> 128
64
> 128
> 128
64
Escherichia coli
ATCC 10536
8
Proteus mirabilis
ATCC 27853
32
64
Klebsiella
128
128
64
64
32
32
128
128
pneumoniae
ATCC 10031
Determined after 24 hours of incubation at 37. for the bacteria. All experiments were run in triplicate. Km = kanamycin sulfate.
Table 3 Antifungal activities of 3-O-acylepicatechin derivatives "5 ± 9), and 3-O-acylcatechin derivatives "18 ± 22). Compounds 1 ± 4, 10 ± 17,
and 23 ± 29 "MIC > 128) not shown.
Test organisms
MIC ꢀmg/mL)
5
6
7
8
9
18
19
20
21
22
A
mpB
275
Candida krusei
IFO 1664
128
128
128
128
128
128
64
> 128
16
> 128
64
128
128
128
128
128
128
64
> 128
16
> 128
64
0.5
Candida lusitaniae
ATCC 42720
> 128
> 128
> 128
> 128
> 128
> 128
0.5
Candida albicans
ATCC 10231
64
16
32
64
8
32
0.125
0.25
Candida tropicalis
32
16
16
32
32
16
IFO 10241
Determined after 24 ± 72 hours of incubation at 28 ± 30. for the fungi. All experiments were run in triplicate. AmpB = Amphotericin B.
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2
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Letter¼ Planta Med 2004; 70: 272±276